N-(phosphonacetyl)-L-aspartate (PALA) is a promising antitumor drug. It inhibits pyrimidine nucleotide biosynthesis at the aspartate transcarbamylase reaction potently and specifically because it resembles the transition state for this reaction. In vivo activity against Lewis lung carcinoma and B16 melanoma is excellent, but the antitumor effect on L1210 and xenografts of human colon cancer in mice is absent or slight. One approach to improving the range and efficiency of antitumor activity is to improve the affinity of transition state analogs for aspartate transcarbamylase by incorporating more electronegative atoms at the site of interaction with a proton donor of the enzyme, and several such analogs are to be made. Another approach is to synthesize transition state analogs of pyrimidine-specific carbamyl-P synthetase, a better target since it is much less active than aspartate transcarbamylase and also because it is the site of feedback regulation for the entire pathway. Several analogs are proposed, all derived from the structure of glutamine, to avoid inhibiting the urea cycle carbamyl-P synthetase. Finally, methods for more efficient large experiments are currently limited by the supply of inhibitor.